1. Field of the Invention
This invention generally relates to separation columns, particularly capillaries useful in capillary zone electrophoresis, including capillary electrochromatography, and more particularly to separation columns that includes a photopolymer component. This photopolymer component may be in the form of a frit.
2. Description of Related Art
Over the past decade, capillary zone electrophoresis (CZE), with its high peak capacity (i.e., the number of peaks separated per unit time), has developed into a powerful and widely used technique for separating ionic species by their electrophoretic mobilities. The lack of selectivity for uncharged analytes in CZE, however, has remained more problematic. Several methods have been developed, such as micellar electrokinetic chromatography (MEKC), to help overcome this problem by providing a pseudostationary phase in which uncharged compounds can be separated. The application of methods such as MEKC is limited because of the restricted number of pseudostationary phases that can be employed in this technique.
With the advent of capillary electrochromatography (CEC), where both chromatographic and electrophoretic transport mechanisms are combined, separation and analysis of mixtures of uncharged analytes can be achieved using low sample volumes with high resolution and efficiency. The increased interest in CEC for analytical applications arises from the large plate numbers and relatively high separation speeds achieved and the wide range of stationary phases (those commonly used in high-performance liquid chromatography) that can be used.
Many groups have reported on the use of slurry or electrokinetic packing methods for the fabrication of electrochromatography capillary columns with typical inner diameters of 75 μm. Reversed-phase capillary columns have been prepared, typically with octadecyl silica (ODS) whose particle diameters are on the order of 1.5 to 5 μm. In both packing techniques, the use of retaining frits at both the inlet and the outlet of the capillary has been required to prevent the chromatographic packing material from exiting the capillary. Although systematic studies regarding the effects of the frits on the performance of such capillaries have not been reported, it is thought that these frits may degrade the efficiencies of these capillary columns.
Alternative approaches have been reported for the preparation of capillary columns that avoid the technical problems of frit fabrication and column preparation associated with slurry and electrokinetic packing. One approach uses bonded stationary phases. Capillary columns prepared in this manner, however, suffer from low retention and low sample capacities as well as long preparation times. An alternative method for the preparation of open tubular capillary columns uses monolithic packing technology. For example, Dulay et al. have described the preparation and characterization of monolithic porous capillary columns loaded with chromatographic particles based on sol-gel chemistry (Dulay et al., Anal. Chem., 70, pp. 5103-5107, 1998).
Nevertheless, where separation columns are desired with packing material requiring frits, it would be desirable to have simple and reproducible procedures for fabricating frits.
The conventional method of frit fabrication for a particle-packed column involves thermal sintering of a section of the packing material, such as ODS (octadecyl silica particles). This approach has several disadvantages, including (1) difficulty in generating the frit reliably and reproducibly, (2) alteration of the characteristics of the stationary phase within the frit itself, (3) difficulty in controlling the porosity of the frit, (4) weakness of the capillary at the location of the frit, (5) band broadening caused by the frit, (6) bubble formation and adsorption of polar analytes on the frit. These problems can directly affect the column performance and column-to-column reproducibility.